Among the heterogeneous population of cells in a tumor is a subset of cells that share characteristics with stem cells and which are termed tumor initiating cells (TICs). TICs exhibit a robust capacity to self-renew and thus play a significant role in tumor initiation. They also exhibit high metastatic potential in vivo, which is manifested in vitro as high motility and invasiveness. Notably, TICs are also resistant to radiation and many currently available chemotherapeutics. Thus, TICs are a novel and therapeutically powerful anti-cancer target. However, little is known their biology, which has hindered the development of TIC-specific chemotherapeutics. We hypothesize that the NF-kB signaling pathway is required for the TIC phenotype in breast cancer cells. This model is supported by data that inhibition of NF-kB in some contexts results in a reduction in cellular self-renewal and NF-kB is involved in the only cellular process to be linked to the generation of TICs, the epithelial-to-mesenchymal transition (EMT). In this proposal, we will test whether NF-kB signaling is preferentially activated in breast cancer TICs and is required for the TIC phenotype in primary human breast tumor samples, human breast cancer cell lines, and murine models of human HER2+ and basal breast cancer. Given the potential association of NF-kB and EMT in TICs, we will also investigate the hypothesis that at least part of the role of NF-kB in TICs is the stimulation of EMT. Finally, we will investigate whether a signaling cascade that depends on the protein kinase TAK1 regulates NF-kB activity in TICs and determine which NF-kB target genes are important for the TIC phenotype. We anticipate that these studies will facilitate the identification of novel chemotherapeutic targets for the treatment of breast cancer. PUBLIC HEALTH RELEVANCE: Breast tumors are thought to contain a sub-set of cancer cells termed tumor initiating cells (TICs) that are responsible for the establishment of primary tumors and promoting metastasis and which are especially resistant to the therapies currently available to patients. Little is currently known about TIC biology, but recent evidence suggests that the NF-kB family of proteins may be important. As such, work in this study to delineate the role of NF-kB in TICs, how NF-kB is regulated in such cells, and how NF-kB promotes the TIC phenotype will facilitate the development of novel chemotherapeutics for breast cancer.